基于白纹伊蚊转录组的溴氰菊酯抗性相关基因分析及验证

Analysis and verification of deltamethrin resistance-related genes based on the transcriptome of Aedes albopictus

目的 通过对白纹伊蚊溴氰菊酯抗性品系(DR)和敏感品系(DS)进行转录组分析,筛选可能与抗性相关的差异表达基因。方法 提取白纹伊蚊实验室敏感品系(DS品系)、溴氰菊酯抗性品系(DR品系)的总RNA进行高通量测序,对差异基因进行COG和KEGG的功能注释与分析,筛选出可能与抗性相关的差异表达基因,并进行实时定量PCR验证。结果 DR品系的LC50为2 087.75μg/L,相较于DS品系,抗性倍数为109.8倍,达到了高度抗性标准。RNA-seq测序后基于FPKM值进行差异性分析结果显示,差异表达的基因共有551个,其中在DR品系中上调表达基因有278个,下调表达基因有273个。COG和KEGG功能富集结果显示,显著性富集的通路和蛋白质功能预测均主要与物质代谢相关。基于此进一步分析了代谢抗性相关基因,且10个目标基因的RNA-seq结果与实时定量PCR结果在DR和DS品系中表达趋势具有一致性。结论 白纹伊蚊可能通过上调表达细胞色素P450基因、谷胱甘肽S转移酶基因、羧酸酯酶、表皮蛋白等基因应对杀虫剂的选择压力。

Objective Transcriptome analysis was performed on deltamethrin-resistant(DR)and deltamethrin-sensitive(DS)strains of Aedes albopictus to screen differentially expressed genes that may be related to resistance.Methods Total RNA was extracted from the susceptible strain(DS)and deltamethrin-resistant strain(DR)of Aedes albopictus for high-throughput sequencing,and the differentially expressed genes were annotated and analyzed by COG(Cluster of Orthologous Groups of proteins)and KEGG(Kyoto Encyclopedia of Genes and Genomes).The differentially expressed genes that might be related to the resistance of Aedes albopictus were screened out and verified by real-time PCR.Results The resistance test results showed that the LCso of DR strain was 2087.75μg/L,109.8 folds of the DS strain,at a high level resistance.The results of differential analysis based on FPKM value after RNA-seq sequencing showed that there were 551 differentially expressed genes in DR strain,278 up-regulated and 273 down-regulated.COG and KEGG functional enrichment results showed that significantly enriched pathways and protein function prediction were mainly related to substance metabolism.Based on this,the metabolic resistance-related genes were further analyzed,and the RNA-seq results of 10 target genes were consistent with the real-time PCR results in DR and DS strain.Conclusions Aedes albopictus can respond to insecticide stress by upregulating the expression of genes as cytochrome P450,glutathione S-transferase,carboxy esterase,cuticular protein.